To investigate the effect of CD300LG-γ induction on the cytotoxic activity of CIK. Eukaryotic expression plasmid hCD300LG-γ/pEGFP-C3, which can express human CD300LG-γ, was constructed and transfected into CHO cells by lipofectamine. The expression of CD300LG- was confirmed by immunofluorescence, RT-PCR, and Western Blot. To produce CIK cells, human peripheral blood mononuclear cells (PBMC) were isolated and induced, respectively, by cell lysates extracted from hCD300LG-γ/CHO cells, pEGFP-C3/CHO cells, and CHO cells, concurrently with the standard CIK inductive agent. The cytotoxic activity of these CIK cells against hCD300LG-γ/CHO cells, pEGFP-C3/CHO cells, CHO cells, and K562 cells was tested. The results showed that eukaryotic expression of plasmid hCD300LG-γ/pEGFP-C3 was constructed and transfected into CHO cells successfully. After being induced by cell lysates, the cytotoxicity of hCD300LG-γ/CHO-CIK was improved compared with the other CIK cells. In particular, the activity of killing pEGFP-C3/CHO and CHO cells was improved significantly. Meanwhile, the activity of hCD300LG-/CHO-CIK killing K562 was improved significantly compared with the other CIK cells. The results indicated that hCD300LG- induction can significantly improve the killing activity of CIK cells.